Synapses are intercellular junctions that mediate neuronal communication, and many synaptic molecules regulate their functions. Recent evidence suggests that cocaine exposure causes abnormalities in synaptic transmission in a minority of neurons that play a causal role in cocaine-induced psychomotor sensitization. A better understanding of cocaine effects on that minority of neurons and their synapse function may provide better treatment strategies for addiction, and thereby might reduce the burden of disease. Towards these ends, identifying behaviorally relevant circuits and then analyzing individual transcript me of neurons associated with those circuits can provide a wealth of information about potential mechanisms of addiction. To achieve such goal, I propose to genetically trace neuronal circuits that are activated by cocaine exposure or context using a FosCreER transgenic mouse line, which allows active neurons to be genetically labeled when the tamoxifen is present. Using FosCreER mouse I will also try to establish the causality of cocaine activated circuits for cocaine related behaviors by blocking activity of those circuits wit virally delivered Cre recombinase dependent expression of the light chain of tetanus toxin. In parallel, I will analyze the transcriptome of activated single neurons in the same circuit by RNAseq and Q-PCR and searching for candidate genes whose expression changes co-vary with altered synaptic properties. In preliminary experiments, I have developed a novel single neuron isolation method using FACS sorting combined with an integrated microfluidic system (C1 Fluidigm). This novel approach enabled me to achieve enhanced purity and efficiency of analyzing single neurons' transcriptome when compared to previous methods. For the independent phase, I will first explore the role of dopamine in cocaine induced gene expression changes. Evidence suggests that cocaine's reinforcing effects depend on its ability to rapidly block the dopamine transporter. To further test if the observed gene expression changes are related to those reinforcing effects, I will test the key gene expression changes in the knock-in mice that have a functional but cocaine-insensitive dopamine transporter. In parallel, I will employ the methods developed in training phase to study cocaine self-administration model. Finally, the role of individual transcriptional changes will be explored by in vitro and in vivo manipulations. The extensive set of experiments in this proposal combines new tools to extract biological insights about the process of cocaine addiction, which will guide the future design of mechanism based targeted pharmacological interventions.